Role of the STAR proteins in early mammalian development [iCLIP]
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ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE153798 | GEO | 2024/08/02
REPOSITORIES: GEO
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Project description:RNA binding proteins (RBPs), are emerging as important master regulators of the biology of stem cells in both pluripotency and differentiation. The Signal transduction and activation of RNA (STAR) is family of proteins shown to be involved in mammalian development. We characterized the role of two members of the family, Sam68 and Quaking (QKI), in pluripotency and differentiation of mouse embryonic stem cells (mESCs) by making use of CRISPR-Cas9 generated knock-out (KO) cell lines along with RNA-sequencing, ribosome profiling and RNA crosslinking followed by immunoprecipitation (iCLIP2). Our results indicate that both proteins are important in the regulation of mESCs cell cycle and self-renewal and are indispensable for proper cardiomyocyte differentiation by regulating expression, alternative splicing and translation of the cardiogenic factors GATA4. This work highlights the importance of the RBPs during the differentiation of mESCs.
2024-08-02 | GSE153799 | GEO
Project description:RNA binding proteins (RBPs), are emerging as important master regulators of the biology of stem cells in both pluripotency and differentiation. The Signal transduction and activation of RNA (STAR) is family of proteins shown to be involved in mammalian development. We characterized the role of two members of the family, Sam68 and Quaking (QKI), in pluripotency and differentiation of mouse embryonic stem cells (mESCs) by making use of CRISPR-Cas9 generated knock-out (KO) cell lines along with RNA-sequencing, ribosome profiling and RNA crosslinking followed by immunoprecipitation (iCLIP2). Our results indicate that both proteins are important in the regulation of mESCs cell cycle and self-renewal and are indispensable for proper cardiomyocyte differentiation by regulating expression, alternative splicing and translation of the cardiogenic factors GATA4. This work highlights the importance of the RBPs during the differentiation of mESCs.
2024-08-01 | GSE153800 | GEO
Project description:The paralog RNA binding proteins (RBPs) Sam68 and SLM2 are co-expressed in the cerebral cortex and display very similar splicing activity. However, their relative function(s) in this context is unknown. By performing a time-course analysis, we found that these RBPs exhibit an opposite expression pattern during development. Sam68 expression declines postnatally while SLM2 increases after birth, and this developmental pattern is reinforced by hierarchical control of Sam68 expression by SLM2. Analysis of Sam68:Slm2 double knockout (Sam68:Slm2dko) mice revealed hundreds of exons that are sensitive to concomitant ablation of these proteins. Moreover, parallel analysis of single and double knockout cortices indicated that exons regulated mainly by SLM2 are characterized by a dynamic splicing pattern during development, whereas Sam68-dependent exons are spliced at relatively constant rates. Dynamic splicing of SLM2-sensitive exons is completely suppressed in the Sam68:Slm2dko developing cortex. Sam68:Slm2dko mice die perinatally with defects in neurogenesis and in neuronal differentiation, and the development of a hydrocephalus, consistent with splicing alterations in genes related to these biological processes. Thus, our study reveals that maintenance of the Sam68 and Slm2 paralog genes encoding homologous RBPs enables the orchestration of a dynamic splicing program while ensuring a robust redundant mechanism that supports proper cortical development.
2024-02-12 | GSE229749 | GEO
Project description:Background: Cellular RNA binding proteins (RBPs) have multiple roles in post-transcriptional control, and some have been shown to bind DNA. However, the global localization and the general chromatin-binding ability of RBPs have not been well-characterized and remain undefined in hematopoietic cells. Results: We first provided a full view of RBPs’ distribution pattern in nucleus and screened for chromatin-enriched RBPs (Che-RBPs) in different human cells. Subsequently, by generating ChIP-seq, CLIP-seq and RNA-seq datasets and conducting combined analysis, the transcriptional regulatory potential of certain hematopoietic Che-RBPs were predicted and from which quaking (QKI5) emerged as a potential transcriptional activator during monocytic differentiation. QKI5 was over-represented in gene promoter regions, independent of RNA or transcription factors. Furthermore, promoter-occupied QKI5 activated the transcription of several critical monocytic differentiation-associated genes, including CXCL2, IL16 and PTPN6. Finally, we showed that the differentiation-promoting activity of QKI5 is largely dependent on CXCL2, irrespective of its RNA-binding capacity. Conclusions: Our study indicates that Che-RBPs are versatile factors that orchestrate gene expression in different cellular contexts, and identifies QKI5, a classic RBP regulating RNA processing, as a novel transcriptional activator during monocytic differentiation.
2023-06-23 | GSE178800 | GEO
Project description:Background: Cellular RNA binding proteins (RBPs) have multiple roles in post-transcriptional control, and some are shown to bind DNA. However, the global localization and the general chromatin-binding ability of RBPs are not well-characterized and remain undefined in hematopoietic cells. Results: We first provide a full view of RBPs' distribution pattern in the nucleus and screen for chromatin-enriched RBPs (Che-RBPs) in different human cells. Subsequently, by generating ChIP-seq, CLIP-seq and RNA-seq datasets and conducting combined analysis, the transcriptional regulatory potential of certain hematopoietic Che-RBPs are predicted. From this analysis, quaking (QKI5) emerges as a potential transcriptional activator during monocytic differentiation. QKI5 is over-represented in gene promoter regions, independent of RNA or transcription factors. Furthermore, DNA-bound QKI5 activates the transcription of several critical monocytic differentiation-associated genes, including CXCL2, IL16 and PTPN6. Finally, we show that the differentiation promoting activity of QKI5 is largely dependent on CXCL2, irrespective of its RNA binding capacity. Conclusions: Our study indicates that Che-RBPs are versatile factors that orchestrate gene expression in different cellular contexts, and identifies QKI5, a classic RBP regulating RNA processing, as a novel transcriptional activator during monocytic differentiation.
2021-09-09 | GSE161943 | GEO
Project description:RNA-binding proteins (RBPs) play integral roles in gene regulation, yet only a small fraction of RBPs has been studied in the context of stem cells. We here apply RNA interference screen for RBPs in mouse embryonic stem cells (ESCs) and identify 16 RBPs involved in pluripotency maintenance. Interestingly, six identified RBPs including Krr1 and Ddx47 are part of a complex called Small Subunit Processome (SSUP) that mediates 18S rRNA biogenesis. The SSUP components are preferentially expressed in stem cells and enhance global translational rate, which is critical to sustain the protein levels of labile pluripotency factors such as Nanog and Esrrb. Furthermore, the SSUP proteins are required for efficient reprogramming of induced pluripotent stem cells. Our study uncovers the role of SSUP and the importance of translational control in stem cell fate decision. R1 cells were transfected with 3 different control siRNAs and 3 different siRNAs targeting Krr1 for 48 hrs. Transcriptome profiles were generated by deep sequencing using illumina HiSeq 2000.
2015-09-24 | E-GEOD-73369 | biostudies-arrayexpress
Project description:Here, we used high-resolution mass spectrometry to identify differentially expressed RNA-binding proteins (RBPs) between TE03 (I3) human embryonic stem cells (hESCs) and human foreskin fibroblasts (HFFs). 2102Ep embryonal carcinoma (EC) cells were used as a reference for pluripotent cells. This data accompanies the manuscript: "Uncovering the RNA-binding protein landscape in the pluripotency network of human embryonic stem cells". Abstract: "Embryonic stem cell (ESC) self-renewal and cell-fate decisions are driven by a broad array of molecular signals. While transcriptional regulators have been extensively studied in human ESCs (hESCs), the extent to which RNA-binding proteins (RBPs) contribute to human pluripotency remains unclear. Here, we carry out a proteome-wide screen and identify 810 proteins that directly bind RNA in hESCs. We reveal that RBPs are preferentially expressed in hESCs and dynamically regulated during exit from pluripotency and early lineage specification. Moreover, we show that nearly 200 RBPs are affected by knockdown of OCT4, a master regulator of pluripotency, several dozen of which are directly bound by this factor. Intriguingly, over 20 percent of the proteins detected in our study are putative DNA- and RNA-binding proteins (DRBPs), among them key transcription factors (TFs). Using fluorescently labeled RNA and seCLIP (single-end enhanced crosslinking and immunoprecipitation) experiments, we discover that the pluripotency-associated STAT3 and OCT4 TFs interact with RNA in hESCs and confirm the direct binding of STAT3 to the conserved NORAD long-noncoding RNA. Taken together, our findings indicate that RBPs have a more widespread role in human pluripotency than previously appreciated, reinforcing the importance of post-transcriptional regulation in stem cell biology".
2023-05-05 | PXD025284 | Pride
Project description:The proteome of undifferentiated human embryonic stem cells (hESCs) was profiled by deep mass spectrometry-based proteomics of whole-cell extracts from suspension cultures of TE03 cells, in four biological replicates. This data accompanies the manuscript: "Uncovering the RNA-binding protein landscape in the pluripotency network of human embryonic stem cells". Abstract: "Embryonic stem cell (ESC) self-renewal and cell-fate decisions are driven by a broad array of molecular signals. While transcriptional regulators have been extensively studied in human ESCs (hESCs), the extent to which RNA-binding proteins (RBPs) contribute to human pluripotency remains unclear. Here, we carry out a proteome-wide screen and identify 810 proteins that directly bind RNA in hESCs. We reveal that RBPs are preferentially expressed in hESCs and dynamically regulated during exit from pluripotency and early lineage specification. Moreover, we show that nearly 200 RBPs are affected by knockdown of OCT4, a master regulator of pluripotency, several dozen of which are directly bound by this factor. Intriguingly, over 20 percent of the proteins detected in our study are putative DNA- and RNA-binding proteins (DRBPs), among them key transcription factors (TFs). Using fluorescently labeled RNA and seCLIP (single-end enhanced crosslinking and immunoprecipitation) experiments, we discover that the pluripotency-associated STAT3 and OCT4 TFs interact with RNA in hESCs and confirm the direct binding of STAT3 to the conserved NORAD long-noncoding RNA. Taken together, our findings indicate that RBPs have a more widespread role in human pluripotency than previously appreciated, reinforcing the importance of post-transcriptional regulation in stem cell biology".
2023-05-05 | PXD025278 | Pride
Project description:Here, we performed RNA-interactome capture (RIC) on nuclear fractions from human embryonic stem cells (hESCs). The poly(A)+ RNA-bound proteome was determined by UV light-mediated cross-linking (CL) of RNAs to proteins in living cells, followed by nuclei isolation, oligo(dT) purification of poly(A)-RNA-protein complexes, and mass spectrometry analysis of captured proteins. As a control, we applied a similar strategy to non-cross-linked (non-CL) samples. RIC was performed in four independent biological replicates. This data accompanies the manuscript: "Uncovering the RNA-binding protein landscape in the pluripotency network of human embryonic stem cells". Abstract: "Embryonic stem cell (ESC) self-renewal and cell-fate decisions are driven by a broad array of molecular signals. While transcriptional regulators have been extensively studied in human ESCs (hESCs), the extent to which RNA-binding proteins (RBPs) contribute to human pluripotency remains unclear. Here, we carry out a proteome-wide screen and identify 810 proteins that directly bind RNA in hESCs. We reveal that RBPs are preferentially expressed in hESCs and dynamically regulated during exit from pluripotency and early lineage specification. Moreover, we show that nearly 200 RBPs are affected by knockdown of OCT4, a master regulator of pluripotency, several dozen of which are directly bound by this factor. Intriguingly, over 20 percent of the proteins detected in our study are putative DNA- and RNA-binding proteins (DRBPs), among them key transcription factors (TFs). Using fluorescently labeled RNA and seCLIP (single-end enhanced crosslinking and immunoprecipitation) experiments, we discover that the pluripotency-associated STAT3 and OCT4 TFs interact with RNA in hESCs and confirm the direct binding of STAT3 to the conserved NORAD long-noncoding RNA. Taken together, our findings indicate that RBPs have a more widespread role in human pluripotency than previously appreciated, reinforcing the importance of post-transcriptional regulation in stem cell biology".
2023-05-05 | PXD025262 | Pride
Project description:Quaking are RNA binding proteins, which are known to regulate the expression of different genes at the post-transcriptional level. Genetic interference with quaking a (qkia) and quaking c (qkic) leads to major myofibril defects during zebrafish development, without affecting early muscle differentiation. In order to understand how qkia and qkic jointly regulate myofibril formation, we performed a comparative analysis of the transcriptome of qkia/qkic (qkia mutant injected with qkic morpholino) versus control embryos. We show that Quaking activity is required for accumulation of the muscle-specific tropomyosin 3 transcript, tpm3.1. Whereas interference with tmp3.1 function disrupts myofibril formation, reintroducing tpm3.1 transcripts into embryos with reduced Quaking activity can restore structured myofibrils. Thus, we identify tropomyosin as an essential component in the process of myofibril formation and as a relay downstream of the regulator proteins Quaking.
2017-09-17 | GSE71573 | GEO